Pseudohypoparathyroidism: A case of hypocalcemia and hypothyroidism diagnosed during the postpartum period.

We describe a 29-year-old Para 1 post-Emergency Lower Segment Caesarean Section (EMLSCS) for fetal distress and Preterm Rupture of the Membrane (PROM) referred by the Obstetric team for persistent bradycardia. She had the typical features of Albright's Hereditary Osteodystrophy (AHO). The laboratory investigation revealed hypocalcemia, hyperphosphatemia with a high Parathyroid hormone (PTH) level and low free Thyroxine 4 (fT4) with high Thyroid Stimulating Hormone (TSH). The patient was diagnosed with Pseudohypoparathyroidism (PHP) Type 1A associated with TSH resistance based on the somatic features of AHO present as well as biochemical and radiological abnormalities.

Cancer-associated fibroblasts regulate keratinocyte cell-cell adhesion via TGF-ß-dependent pathways in genotype-specific oral cancer.

The interrelationship between malignant epithelium and the underlying stroma is of fundamental importance in tumour development and progression. In the present study, we used cancer-associated fibroblasts (CAFs) derived from genetically unstable oral squamous cell carcinomas (GU-OSCC), tumours that are characterized by the loss of genes such as TP53 and p16INK4A and with extensive loss of heterozygosity, together with CAFs from their more genetically stable (GS) counterparts that have wild-type TP53 and p16INK4A and minimal loss of heterozygosity (GS-OSCC). Using a systems biology approach to interpret the genome-wide transcriptional profile of the CAFs, we show that transforming growth factor-ß (TGF-ß) family members not only had biological relevance in silico but also distinguished GU-OSCC-derived CAFs from GS-OSCC CAFs and fibroblasts from normal oral mucosa. In view of the close association between TGF-ß family members, we examined the expression of TGF-ß1 and TGF-ß2 in the different fibroblast subtypes and showed increased levels of active TGF-ß1 and TGF-ß2 in CAFs from GU-OSCC. CAFs from GU-OSCC, but not GS-OSCC or normal fibroblasts, induced epithelial-mesenchymal transition and down-regulated a broad spectrum of cell adhesion molecules resulting in epithelial dis-cohesion and invasion of target keratinocytes in vitro in a TGF-ß-dependent manner. The results demonstrate that the TGF-ß family of cytokines secreted by CAFs derived from genotype-specific oral cancer (GU-OSCC) promote, at least in part, the malignant phenotype by weakening intercellular epithelial adhesion.

Resection of a physeal bar with complete transverse osteotomy at the metaphysis and Ilizarov external fixation.

The traditional techniques involving an oblique tunnel or triangular wedge resection to approach a central or mixed-type physeal bar are hindered by poor visualisation of the bar. This may be overcome by a complete transverse osteotomy at the metaphysis near the growth plate or a direct vertical approach to the bar. Ilizarov external fixation using small wires allows firm fixation of the short physis-bearing fragment, and can also correct an associated angular deformity and permit limb lengthening. We accurately approached and successfully excised ten central- or mixed-type bars; six in the distal femur, two in the proximal tibia and two in the distal tibia, without damaging the uninvolved physis, and corrected the associated angular deformity and leg-length discrepancy. Callus formation was slightly delayed because of periosteal elevation and stretching during resection of the bar. The resultant resection of the bar was satisfactory in seven patients and fair in three as assessed using a by a modified Williamson-Staheli classification.

Penicillium marneffei infection in a lung transplant recipient.

Penicillium marneffei is a thermally dimorphic fungus that can cause severe opportunistic infections in endemic regions of Southeast Asia, particularly in individuals infected with human immunodeficiency virus-1, but has rarely been reported in solid organ transplant recipients. Herein, we report the first case, to our knowledge, of P. marneffei infection in a lung transplant recipient, occurring in a 41-year-old woman 28 months post lung transplantation, after recent travel to Vietnam. We have reviewed the literature to derive some management principles for this rare infection in this clinical context. The number of P. marneffei infections in transplant recipients may increase, as a result of increasing rates of transplantation and travel to endemic areas.

Oligomer adsorption on dry and wet collagen surfaces.

The development of new biodegradable polymeric tissue adhesives has been almost stagnant for the past 10 years, primarily due to the inability to overcome the problem of inadequate adhesion properties. Efforts at the synthesis and modification of chemical structures by incorporating functional groups have proven futile. This study proposes using simulation as a preliminary move to obtain a better understanding of adsorption behavior on biological tissues. It is hoped that this understanding will subsequently serve as a guide for better polymer design and synthesis. Adsorption under both dry and wet conditions were simulated applying classical molecular mechanics and dynamics (MM/MD) because of their relevancy and efficiency. Twelve types of oligomers and a model collagen surface were constructed, followed by structural optimization and equilibration treatments. The COMPASS force field was used to describe the molecular potential energy surfaces. One strand of the oligomer was then located on top of the collagen surface and their interactions at equilibrium, in terms of van der Waals (vdW) and electrostatic energies, monitored over time. For the wet environment a thick water layer was constructed and placed on top of the oligomer and collagen surface. The results showed that the vdW component dominated physical adsorption for all oligomers, under both dry and wet conditions. This implies that interactions of polymers with tissue surfaces are inherently weak. Functional groups on oligomers could improve adhesion via electrostatic interaction. This interaction is, however, screened off in a wet environment, resulting in a reduction in the adsorption energy for all molecules studied. Of all the oligomers studied, poly(glycine) showed the strongest adsorption to collagen in both dry and wet conditions. Therefore, it is proposed to include the functional groups present in poly(glycine) in future tissue adhesive systems.

Interaction force measurements for the design of tissue adhesives.

Synthesis of tissue adhesives had been carried out in various laboratories in the past decades but the development is currently stalled. One of the key reasons, it is believed, is that researchers have not fully understood and resolved the role of the functional groups that are responsible for good adhesion to biological tissues. Further progress in synthesis is significantly hindered without this fundamental understanding. With this aim in mind, atomic force microscopy (AFM) has been exploited in this work to study the interactions between functional groups that are common to biological tissues. In this work, the AFM tip and substrates were functionalized and used to measure the non-specific interaction among these common functional groups. The ultimate aim of the study is to calculate the interaction force between a single pair of functional groups. A novel calculation method based on the AFM data and probe geometry is presented. The results provide insights into the strength of the bond between different functional groups and the could serve as a guide in selecting the appropriate functional groups in tissue adhesive synthesis. This method could be further applied to studies involving interfaces of biomedical devices where intermolecular interactions are of concern.

Penile metastasis: an unusual presentation of metastatic colonic cancer.

We report here an interesting presentation of a primary colonic carcinoma in a urological setting. A previously unknown case of colonic carcinoma presented with a lesion in the glans penis which was later diagnosed as a secondary deposit from colonic cancer. Penile involvement has been implicated as a metastatic site in several tumours. Although uncommon, this presentation is not unknown. A literature review of this unusual presentation has been performed and is summarised in the article

Further identification and characterization of novel intermediate and mature cleavage products released from the ORF 1b region of the avian coronavirus infectious bronchitis virus 1a/1b polyprotein.

The coronavirus 3C-like proteinase is one of the viral proteinases responsible for processing of the 1a and 1a/1b polyproteins to multiple mature products. In cells infected with avian coronavirus infectious bronchitis virus (IBV), three proteins of 100, 39, and 35 kDa, respectively, were previously identified as mature cleavage products released from the 1b region of the 1a/1b polyprotein by the 3C-like proteinase. In this report, we show the identification of two more cleavage products of 68 and 58 kDa released from the same region of the polyprotein. In addition, two stable intermediate cleavage products with molecular masses of 160 and 132 kDa, respectively, were identified in IBV-infected cells. The 160-kDa protein was shown to be an intermediate cleavage product covering the 100- and 68-kDa proteins, and the 132-kDa protein to be an intermediate cleavage product covering the 58-, 39-, and 35-kDa proteins. Immunofluorescent staining of IBV-infected cells and cells expressing individual cleavage products showed that the 100-, 68-, and 58-kDa proteins were associated with the membranes of the endoplasmic reticulum, and the 39- and 35-kDa proteins displayed diffuse distribution patterns.

The missing link in coronavirus assembly. Retention of the avian coronavirus infectious bronchitis virus envelope protein in the pre-Golgi compartments and physical interaction between the envelope and membrane proteins.

One missing link in the coronavirus assembly is the physical interaction between two crucial structural proteins, the membrane (M) and envelope (E) proteins. In this study, we demonstrate that the coronavirus infectious bronchitis virus E can physically interact, via a putative peripheral domain, with M. Deletion of this domain resulted in a drastic reduction in the incorporation of M into virus-like particles. Immunofluorescent staining of cells coexpressing M and E supports that E interacts with M and relocates M to the same subcellular compartments that E resides in. E was retained in the pre-Golgi membranes, prior to being translocated to the Golgi apparatus and the secretory vesicles; M was observed to exhibit similar localization and translocation profiles as E when coexpressed with E. Deletion studies identified the C-terminal 6-residue RDKLYS as the endoplasmic reticulum retention signal of E, and site-directed mutagenesis of the -4 lysine residue to glutamine resulted in the accumulation of E in the Golgi apparatus. The third domain of E that plays a crucial role in virus budding is a putative transmembrane domain present at the N-terminal region, because deletion of the domain resulted in a free distribution of the mutant protein and in dysfunctional viral assembly.

Low-bit-rate video coding using dense motion field and uncovered background prediction.

It is well known that accurate dense motion field can improve the video coding efficiency. This paper presents a novel Markov random field (MRF) model that estimates both the dense motion and uncovered background fields in image sequences, and the application of these estimates in H.263-based video coding framework.

Identification of a novel cleavage activity of the first papain-like proteinase domain encoded by open reading frame 1a of the coronavirus Avian infectious bronchitis virus and characterization of the cleavage products.

The coronavirus Avian infectious bronchitis virus (IBV) employs polyprotein processing as a strategy to express its gene products. Previously we identified the first cleavage event as proteolysis at the Gly(673)-Gly(674) dipeptide bond mediated by the first papain-like proteinase domain (PLPD-1) to release an 87-kDa mature protein. In this report, we demonstrate a novel cleavage activity of PLPD-1. Expression, deletion, and mutagenesis studies showed that the product encoded between nucleotides 2548 and 8865 was further cleaved by PLPD-1 at the Gly(2265)-Gly(2266) dipeptide bond to release an N-terminal 195-kDa and a C-terminal 41-kDa cleavage product. Characterization of the cleavage activity revealed that the proteinase is active on this scissile bond when expressed in vitro in rabbit reticulocyte lysates and can act on the same substrate in trans when expressed in intact cells. Both the N- and C-terminal cleavage products were detected in virus-infected cells and were found to be physically associated. Glycosidase digestion and site-directed mutagenesis studies of the 41-kDa protein demonstrated that it is modified by N-linked glycosylation at the Asn(2313) residue encoded by nucleotides 7465 to 7467. By using a region-specific antiserum raised against the IBV sequence encoded by nucleotides 8865 to 9786, we also demonstrated that a 33-kDa protein, representing the 3C-like proteinase (3CLP), was specifically immunoprecipitated from the virus-infected cells. Site-directed mutagenesis and expression studies showed that a previously predicted cleavage site (Q(2583)-G(2584)) located within the 41-kDa protein-encoding region was not utilized by 3CLP, supporting the conclusion that the 41-kDa protein is a mature viral product.

Long-term outcome and disease control in near-fatal asthma.

Patients who have survived an episode of intubation and mechanical ventilation for acute respiratory failure due to a severe and unresponsive asthmatic attack are considered to have experienced a near-fatal asthma (NFA) attack. Such patients are at a higher risk of similar severe attacks and hence of death in the future. The aims of the study were to: (i) evaluate the outcome; (ii) identify any persistent deficiencies in asthma management, and (iii) assess self-management knowledge in survivors of NFA. Ninety-three consecutive patients who had been treated for NFA in the Intensive Care Unit of an urban teaching hospital in Singapore from 1992 to 1997 were studied. All hospital records were reviewed retrospectively. Survivors were then invited to attend a questionnaire interview and to have lung function tests performed. Of the original cohort (OC) of 93 patients with NFA (mean age 55.2 years), 18 (19% OC) patients (mean age 64 years) had died while in hospital and 75 (81% OC) patients survived the initial episode of NFA and were discharged home (DH). The long-term outcome of this DH group was: 13 patients had died (17% DH) and 62 (83% DH) survived. Of these survivors, 35 were interviewed while 27 declined or were not contactable. This interview yielded the following information: (i) Hospitalisation in the past year: 66% had no hospital admission; of the 31.4% who had 2 or more admissions, most had a further NFA attack. (ii) Health care: The majority of patients (71.4%) were monitored by a single doctor. (iii) Patient knowledge of disease management was deemed good to fair for trigger avoidance (77%), for appropriate drug usage (97%). (iv) Satisfactory inhaler skill (80%). NFA is associated with a high intrahospital and long-term mortality. Although most survivors of NFA appeared to have satisfactory care and a fair understanding of medication usage, a significant minority continue to pose much morbidity and risk for death.

Characterisation of a papain-like proteinase domain encoded by ORF1a of the coronavirus IBV and determination of the C-terminal cleavage site of an 87 kDa protein.

Our previous studies have shown that two overlapping papain-like proteinase domains (PLPDs) encoded by the IBV sequence from nucleotides 4155 to 5550 is responsible for cleavage of the ORF 1a polyprotein to an 87 kDa protein. In this study, we demonstrate that only the more 5' one of the two domains, PLPD-1 encoded between nucleotides 4155 and 5031, is required for processing to the 87 kDa protein. Site-directed mutagenesis studies have shown that the Cys1274 and His1435 residues are essential for the PLPD-1 activity, suggesting that they may be the components of the catalytic centre of this proteinase. Coexpression and immunoprecipitation studies have further revealed that PLPD can interact with the 87 kDa protein. Meanwhile, data obtained from the construction and expression of a series of deletion mutants have indicated that the 87 kDa protein is encoded by the 5'-most 2600 bp part of ORF1a. further deletion and mutagenesis studies are underway to determine precisely the C-terminal cleavage site of the 87 kDa protein.

Regulation of mRNA 1 expression by the 5'-untranslated region (5'-UTR) of the coronavirus infectious bronchitis virus (IBV).

In this report, we show that expression of the coronavirus IBV mRNA1 is regulated by its 5'-UTR. Evidence presented demonstrates that the IBV sequence from nucleotide 1 to 1904 directs very inefficient synthesis of a product of approximately 43 kDa. Deletion of either the first 362 bp or the whole part of the 5'-UTR, however, dramatically increased the expression of the 43 kDa protein species. The mechanisms involved were investigated by two different approaches. Firstly, translation of the same construct in the presence of [3H]-leucine ruled out the possibility that initiation of small reading frames from non-AUG codons located in the 5'-UTR may compete with the authentic AUG initiation codon, and therefore inhibit the expression of ORF 1a. Secondly, expression and deletion analyses of a dicistronic construct showed that translation of the 43 kDa protein was initiated by ribosome internal entry mechanism. These studies suggest that a 'weak' ribosome internal entry signal is located in the 5'-UTR and is involved in the regulation of mRNA1 expression.

Characterization of the two overlapping papain-like proteinase domains encoded in gene 1 of the coronavirus infectious bronchitis virus and determination of the C-terminal cleavage site of an 87-kDa protein.

In a previous report, we showed that proteolytic processing of an 87-kDa mature viral protein from the coronavirus infectious bronchitis virus (IBV) 1a and 1a/1b polyproteins was mediated by two putative overlapping papain-like proteinase domains (PLPDs) encoded within the region from nucleotides 4243 to 5553 of ORF 1a (Liu et al., 1995). In this study, we demonstrate that only the first domain, PLPD-1, is responsible for this cleavage, as deletion of the second domain did not affect the formation of the 87-kDa protein. Site-directed mutagenesis studies further showed that a previously predicted nucleophilic cysteine residue (Cys1274) and a histidine residue (His1437) were essential for the proteinase activity, indicating that they may be important components of the catalytic center of the proteinase. Meanwhile, expression of a series of deletion mutants revealed that the 87-kDa protein was encoded by the 5'-most 2.6 kb of ORF 1a. Deletion and amino acid substitution mutation studies demonstrated that the Gly673-Gly674 dipeptide bond was most likely the cleavage site responsible for releasing the C-terminus of the 87-kDa protein from the 1a and 1a/1b polyproteins.

Differences in the hydrolysis of lactose and other substrates by beta-D-galactosidase from Kluyveromyces lactis.

The hydrolysis of o-nitrophenyl galactopyranoside and lactose by beta-D-galactosidase from Kluyveromyces lactis was enhanced by the addition of Mg2+ and Mn2+, but the rates of activation by each metal on both substrates were not the same. The Co2+, Zn2+, and Ni2+ activated the o-nitrophenyl galactopyranoside-hydrolyzing activity of the enzyme, but these same metals inhibited the lactose-hydrolyzing activity. The addition of Mg2+ and EDTA to the assay buffer increased the hydrolysis of o-nitrophenyl galactopyranoside and lactose at different rates. The responses of o-nitrophenyl galactopyranoside and lactose to the enzyme activity were different as a function of pH. The hydrolyzing activity toward both substrates also was influenced by the concentration of the phosphate in the assay buffer. However, the profile of the enzyme activity toward each substrate was different as a function of concentration. Because the assay of beta-galactosidase using o-nitrophenyl galactopyranoside is fast and convenient, the estimation of lactose-hydrolyzing activity of the enzyme has frequently been made based on the assay of o-nitrophenyl galactopyranoside hydrolysis. As shown in this study, a slight change in the conditions of the assay system and the enzyme application may cause changes in the ability of the enzyme to hydrolyze both lactose and o-nitrophenyl galactopyranoside. The change in o-nitrophenyl galactopyranoside-hydrolyzing activity is not always consistent with that of the lactose-hydrolyzing activity under the given condition, which may cause an inaccurate estimation of the enzyme activity in the enzyme preparation as well as in actual applications of the enzyme.

Amplification and sequencing of varicella-zoster virus (VZV) gene 4: point mutation in a VZV strain causing chickenpox during pregnancy.

The varicella-zoster virus (VZV) causes chickenpox (varicella) as the primary disease and shingles (zoster) as a recurrent manifestation of infection, both being generally benign and self-limiting. While these infections may be severe in adults and even life-threatening in immunosuppressed individuals, they may be amenable to effective antiviral drugs or varicella-zoster immune globulin, provided the treatment is administered early. The prompt diagnosis of VZV infections may be accelerated by rapid, sensitive and specific molecular techniques such as amplification by polymerase chain reaction (PCR) compared with slower and more cumbersome tissue culture and serological procedures. Based on the VZV gene 4 which encodes a transcriptional activator, primers were designed for use in PCR to amplify a target fragment of 381 bp. Distinct diagnostic bands were observed by agarose gel electrophoresis of PCR products of VZV strains isolated from 11 varicella and 7 zoster patients in Singapore, as well as of the Japanese vaccine Oka strain. The detection sensitivity of this PCR assay was determined to be 1 pg of purified VZV DNA equivalent to about 7,000 viral DNA copies. No target bands were amplified from negative control templates from five related human herpes-viruses and from human DNA. The specificity of the PCR products was ensured by direct cycle DNA sequencing, which revealed complete identity of the 18 VZV isolates with the published European Dumas strain. The strong sequence conservation of the target fragment renders this PCR assay highly reliable for detecting the VZV sequence. Only one VZV strain isolated from a patient with varicella during pregnancy exhibited a GGA to GAA point mutation at codon 46 of gene 4, culminating in the non-conservative substitution of Ser with Phe. The predicted secondary structure of the mutant polypeptide portrayed a radical alteration, which may influence its function in transcriptional activation.

Low frequency of endoscopic esophagitis in Asian patients.

The aim of this study was to determine the frequency of endoscopic esophagitis in patients seen for upper gastrointestinal complaints in an Asian center. We studied a consecutive series of 11,943 patients undergoing diagnostic esophagogastroduodenoscopy at our unit over a 10-year period. Three hundred and eighty-nine patients (3.3%) had endoscopic esophagitis with no other significant lesion (primary esophagitis), whereas 143 (1.2%) had esophagitis associated with peptic ulcer or gastric or duodenal malignancy (secondary esophagitis). In contrast, peptic ulcer was diagnosed in 2,787 patients (23.3%) and gastric carcinoma in 286 (2.4%). The reported frequency of endoscopic esophagitis among patients undergoing endoscopy in Western countries varied from 9 to 23%. Our data therefore show that endoscopic esophagitis is much less common in Singaporean patients.